Abstract Autoimmune uveitis (AU) is a major cause of visual disability worldwide. Clinical treatment currently involves the use of immunosuppressive drugs, but serious side effects limit their long-term application. In this sense, safer and more effective therapies for AU are needed. The cytotoxic destruction of the retina and related tissues by T helper 1 (Th1) and Th17 cells is the key pathological feature of AU, hence the design of efficient cause-addressing AU treatments should target silencing of Th1/Th17 cells. One therapeutic strategy to suppress the auto-reactive T cells is the use of mesenchymal stem cells (MSCs) as their immune suppressive abilities have been extensively shown in cultures, animal models and patients. However, large variations in MSCs, due to the differences in donors, culture conditions, and tissue sources impede developing of robust MSC therapy. Lately, MSC-derived extracellular vesicles (MSC-EVs) have become an emerging alternative to MSC therapy, as they recapitulate to a large extent the broad therapeutic effects previously attributed to MSCs. Also, EVs display intrinsic cell targeting-properties, stability in circulation without loss of function, and superior safety profile of a cell-free treatment, suggesting MSC-EVs have multiple advantages over cell treatment. Importantly, we recently showed that administration of MSC-EVs prevent the onset of autoimmune disease in murine models, type 1 diabetes, experimental AU (EAU), and Sjogren's syndrome. Therefore, the goal of this proposal is to develop a safe and effective MSC-EV therapy for AU. The overall objective is to define the underlying mechanism by which MSC-EVs suppress autoimmunity and develop strategies to maximize their therapeutic efficacy. To achieve our objective, Aim 1 will identify the therapeutic factors in EVs that are responsible for the MSC-EV mediated action in Au, defining the underlying mechanism by which MSC-EVs suppress Th1/Th17 cells. Aim 2 will develop strategies to maximize the immune suppressive effect of MSC- EVs. Aim 3 will develop strategies to enhance MSC-EV delivery to the target cells. The success of the proposal research will provide essential insights and valuable information for the rational design of EV-based interventions, ultimately leading to a development of the novel, safe and clinically feasible therapy with MSC- EVs for AU treatment. !